LOCUS       sequence-206040-        5931 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for bacterial expression of a His-tagged Y172M mutant of GST
            encoded by preferred codons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5931)
  AUTHORS   Josse L, Sampson CDD, Tuite MF, Howland K, von der Haar T
  TITLE     Codon-Dependent Translational Accuracy Controls Protein Quality in 
            Escherichia coli but not in Saccharomyces cerevisiae
  JOURNAL   bioRxiv 200006
REFERENCE   2  (bases 1 to 5931)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            200006"
COMMENT     Sequence Label: pTH859-gst-optec Y172M
FEATURES             Location/Qualifiers
     source          1..5931
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     50..69
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     153..172
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        153..171
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    172..196
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             211..233
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             246..269
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             948..965
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1018..1036)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1032..1079
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1116..1571
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1203..1222)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1413..1434
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1663..2478)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1663..2478)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2386..2405
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2470..2489
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2600..3188
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3089..3108
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3342..3359
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3374..3513
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3439..3458)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3599..3621
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3615..3806)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4615..5697)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5659..5678
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5698..5775)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg
       61 cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg atgcgtccgg
      121 cgtagaggat cgagatctcg atcccgcgaa attaatacga ctcactatag gggaattgtg
      181 agcggataac aattcccctc tagaaataat tttgtttaac tttaagaagg agatatacca
      241 tgggccacca tcaccatcac catcaccata gcccgattct gggctattgg aaaattaaag
      301 gcctggtgca gccgacccgc ctgctgctgg aatatctgga agaaaaatat gaagaacatc
      361 tgtatgaacg cgatgaaggc gataaatggc gcaacaaaaa atttgaactg ggcctggaat
      421 ttccgaacct gccgtattat attgatggcg atgtgaaact gacccagagc atggcgatta
      481 ttcgctatat tgcggataaa cataacatgc tgggcggctg cccgaaagaa cgcgcggaaa
      541 ttagcatgct ggaaggcgcg gtgctggata ttcgctatgg cgtgagccgc attgcgtata
      601 gcaaagattt tgaaaccctg aaagtggatt ttctgagcaa actgccggaa atgctgaaaa
      661 tgtttgaaga tcgcctgtgc cataaaacct atctgaacgg cgatcatgtg acccatccgg
      721 attttatgct gtatgatgcg ctggatgtgg tgctgatgat ggatccgatg tgcctggatg
      781 cgtttccgaa actggtgtgc tttaaaaaac gcattgaagc gattccgcag attgataaat
      841 atctgaaaag cagcaaatat attgcgtggc cgctgcaggg ctggcaggcg acctttggcg
      901 gcggcgatca tccgccgaaa taataaaagc ttgcggccgc actcgagcac caccaccacc
      961 accactgaga tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg
     1021 ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc
     1081 tgaaaggagg aactatatcc ggattggcga atgggacgcg ccctgtagcg gcgcattaag
     1141 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     1201 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     1261 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     1321 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     1381 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     1441 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     1501 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     1561 gcttacaatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     1621 ttctaaatac attcaaatat gtatccgctc atgaattaat tcttagaaaa actcatcgag
     1681 catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag
     1741 ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg
     1801 gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc
     1861 aaaaataagg ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg
     1921 caaaagttta tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc
     1981 aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa
     2041 tacgcgatcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa
     2101 cactgccagc gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa
     2161 tgctgttttc ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa
     2221 atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc
     2281 tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg
     2341 cttcccatac aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt
     2401 atacccatat aaatcagcat ccatgttgga atttaatcgc ggcctagagc aagacgtttc
     2461 ccgttgaata tggctcataa caccccttgt attactgttt atgtaagcag acagttttat
     2521 tgttcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     2581 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     2641 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     2701 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     2761 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     2821 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     2881 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     2941 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     3001 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     3061 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     3121 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     3181 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     3241 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     3301 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     3361 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     3421 gcaatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact
     3481 ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac acccgctgac
     3541 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     3601 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg
     3661 taaagctcat cagcgtggtc gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc
     3721 agctcgttga gtttctccag aagcgttaat gtctggcttc tgataaagcg ggccatgtta
     3781 agggcggttt tttcctgttt ggtcactgat gcctccgtgt aagggggatt tctgttcatg
     3841 ggggtaatga taccgatgaa acgagagagg atgctcacga tacgggttac tgatgatgaa
     3901 catgcccggt tactggaacg ttgtgagggt aaacaactgg cggtatggat gcggcgggac
     3961 cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta atacagatgt aggtgttcca
     4021 cagggtagcc agcagcatcc tgcgatgcag atccggaaca taatggtgca gggcgctgac
     4081 ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag
     4141 gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc
     4201 tgctaaccag taaggcaacc ccgccagcct agccgggtcc tcaacgacag gagcacgatc
     4261 atgcgcaccc gtggggccgc catgccggcg ataatggcct gcttctcgcc gaaacgtttg
     4321 gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca agattccgaa taccgcaagc
     4381 gacaggccga tcatcgtcgc gctccagcga aagcggtcct cgccgaaaat gacccagagc
     4441 gctgccggca cctgtcctac gagttgcatg ataaagaaga cagtcataag tgcggcgacg
     4501 atagtcatgc cccgcgccca ccggaaggag ctgactgggt tgaaggctct caagggcatc
     4561 ggtcgagatc ccggtgccta atgagtgagc taacttacat taattgcgtt gcgctcactg
     4621 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     4681 gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg
     4741 caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct
     4801 ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga
     4861 gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc gcagcccgga
     4921 ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt
     4981 gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca
     5041 gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc
     5101 agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg
     5161 gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat
     5221 aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca
     5281 ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact
     5341 gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc
     5401 taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac
     5461 aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg
     5521 tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc
     5581 ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac
     5641 ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt
     5701 caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg
     5761 ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc
     5821 agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg
     5881 agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg c
//
